The Cytotoxic Cell Studies Group has made significant advances in understanding the molecular mechanism of natural killer (NK) cell function by examining the role of a novel gene, Nktr, in the tumor recognition/killing mechanism of NK cells. The crucial role of the NK-TR protein in target cell recognition/killing was demonstrated by inhibiting NK-TR expression in primary human and mouse NK cells with a retroviral vector producing antisense Nktr RNA. Experiments with freshly isolated human and mouse NK cells have demonstrated a dramatic loss of NK activity relative to control retrovirus infections. The possible involvement of Nktr in RNA splicing has been strengthened by identification of several Nktr associated proteins using a yeast two- hybrid screen. A protein thought to function as an alternate splicing factor, U2AF-RS2, has been found to specifically associate with the Nktr. Two novel human proteins related to splicing factors have also been shown to interact with the Nktr in a specific manner. Our current studies are directed towards defining Nktr function in cytotoxic cells, and developing gene knockout mice. Mice heterozygous for an Nktr null mutation have been generated, and are currently being bred to produce homozygous Nktr-deficient mice. We are continuing our study of the Ly49 family expressed on murine NK cells. The Ly49-A,C, and G family members are inhibitory receptors that recognize MHC class I. We have shown that Ly-49D represents an activating receptor. Current studies are focussed on defining the signal transduction molecules that associate with the Ly49D receptor.